Food allergen testing: should you use ELISA, PCR, or LFA methods?

Food allergen testing plays a crucial role in ensuring the safety and well-being of individuals with food allergies. Allergens are proteins present in certain foods that can trigger adverse reactions in susceptible individuals, ranging from mild symptoms to life-threatening anaphylaxis. Accurate allergen detection is essential to prevent allergen cross-contamination, mislabeling, and unintended exposure in food products. In recent years, several methods have been developed to accurately detect and quantify allergens in food products and food-processing facilities. The most widely used technologies in this field are Enzyme-linked Immunosorbent Assay (ELISA), Polymerase Chain Reaction (PCR), and Lateral Flow Assays (LFA). Each of these methods offers distinct advantages and applications in food allergen detection. Collectively, they enable the food industry to effectively detect and quantify allergens, thereby mitigating risks and upholding food safety standards. This blog post explores the differences and applications of these technologies in food allergen detection to emphasize their significance in safeguarding public health and promoting consumer trust in the food supply chain.

Enzyme-linked Immunosorbent Assay (ELISA)

ELISA is a widely used and established technology for food allergen detection. It is based on the principle of antigen-antibody interaction, using antibodies that specifically bind to allergenic proteins present in food samples. The process begins with the preparation of food samples, which usually involves a procedure to extract proteins. These extracts are then added to wells of a microplate that have been coated with antibodies specific to the allergen of interest. If the allergenic protein is present, it binds to these antibodies. A secondary antibody, linked to an enzyme, is then added, binding to the allergen-antibody complex. When a substrate of that enzyme is added, it reacts to produce a measurable signal, typically a color change. The intensity of this signal, measured using a spectrophotometer, correlates with the amount of allergen present, allowing for precise quantification. ELISA’s high sensitivity and specificity enable it to detect and quantify allergens at very low concentrations, which is crucial for food testing laboratories. The method is versatile, capable of testing a wide range of food products, from raw ingredients to processed foods.

Applications:

• Quality control in food manufacturing to prevent cross-contamination of allergens.
• Regulatory compliance to ensure accurate labeling of allergens on food products.
• Analysis of egg and milk allergens as qPCR is not able to differentiate between the egg and milk from chicken and cow, respectively.
• ELISA is the standard method for gluten analysis as indicated by the Codex Alimentarius.
• ELISA is the preferred method when a quantitative answer is required.

Polymerase Chain Reaction (PCR)

PCR is a molecular biology technique that amplifies specific DNA sequences, making it a powerful tool in food allergen detection. By targeting DNA sequences unique to allergenic proteins or species, PCR can accurately identify the presence of allergens, even in processed or heavily treated food products. The process begins with the extraction of DNA from food samples. This DNA is then mixed with primers, which are short nucleotides that hybridize specifically with the target allergen’s DNA. The mixture is placed in a thermal cycler, which repeatedly heats and cools the sample to promote the synthesis of new DNA strands. This cycle of heating and cooling is repeated multiple times, exponentially amplifying the target DNA sequence. The amplified DNA is then detected by real-time PCR, which provides a measurable signal indicating the presence of the allergen. PCR is recognized by its multiplexing capability and offers a higher throughput than other techniques for allergen testing.

Applications:

• Detection of allergens in complex food matrices, such as baked goods, where protein structure may be altered.
• Verification of allergen-free claims in food products, especially in products with multiple ingredients.
• Analysis of celery and fish allergens as celery is a low-protein matrix with many cross-reactivities and there is no common antigen for fish detection using ELISA.
• Analysis of highly processed foods, including hydrolyzed and fermented samples, as DNA is a more stable molecule.
• PCR is the preferred method when a qualitative answer is required.

Lateral Flow Assays (LFA)

Lateral flow assays (LFAs) are a popular and efficient technology for food allergen detection, known for their simplicity and rapid results. These devices use the principle of immunochromatography, where antibodies are used to detect the presence of allergenic proteins in a variety of sample types. The sample is applied to the sample pad of the lateral flow device which is comes in a strip format. The sample migrates along the test strip by capillary action. As it moves, it encounters a conjugate pad containing antibodies that are specific to the allergen of interest and are labeled with a detectable marker, such as colloidal gold. If the allergenic protein is present, it binds to these labeled antibodies. The sample continues to migrate along the strip to the test line, where another set of antibodies specific to the allergen is immobilized. The allergen-antibody complex binds to these immobilized antibodies, forming a visible line. The intensity of this line indicates the presence and, to some extent, the quantity of the allergen. A control line further along the strip confirms that the test has worked correctly. LFAs are highly valued for their rapid results, often providing answers within minutes. They are user-friendly and do not require specialized equipment or extensive training, making them suitable for on-site testing in various settings, including food manufacturing facilities and retail environments.

Applications:

• On-site allergen testing for food manufacturers, allowing them to verify the absence of allergens in facilities, including equipment and utensils.
• Screening of incoming raw materials in food production facilities to prevent allergen contamination.
• LFAs are suitable for point-of-care settings, including commercial kitchens, providing quick and reliable allergen testing to ensure consumer safety.
• Validation and monitoring of hygiene and sanitation processes by analyzing environmental swabs and rinse waters.

Combining the methods for enhanced results

Often, using a combination of the different technologies can enhance food allergen testing by providing complementary strengths and overcoming limitations of individual methods. Here are some scenarios where an integrated approach can lead to more reliable results:

1. Complex Food Matrices:

   In food products with complex matrices or heavily processed ingredients, a combination of PCR and ELISA can be beneficial. PCR can target and amplify specific DNA sequences of allergenic species that may be degraded or denatured during processing, while ELISA can provide quantitative information on the actual protein levels present in the sample. By combining the sensitivity of PCR with the specificity of ELISA, a more comprehensive analysis of allergen content in such complex food matrices can be achieved.
   
   

2. On-Site Rapid Testing:

   In situations where quick on-site allergen testing is required, a combination of LFAs and PCR can offer a rapid and reliable solution. LFAs provide rapid qualitative results that can be easily interpreted without the need for specialized equipment, while PCR can confirm the presence of allergens at a molecular level.
   
   

3. Allergen Label Verification:

   When verifying allergen-free claims in food products, an integrated approach involving ELISA and LFAs can enhance the reliability of results. ELISA can provide accurate quantification of allergens, while LFAs can offer a quick screening tool for preliminary assessment.

What Invitek Diagnostics has to offer

Are you in need of advanced solutions for food allergen testing? Look no further! We are proud to offer a comprehensive portfolio of all the above-mentioned technologies through our InviScreen®, InviRapid® and InviLisa® product lines.

Our integrated approach to allergen testing enables you to enhance the safety and quality of the food industry by leveraging the combined strengths of these cutting-edge technologies. The infographic below identifies critical points and areas throughout the entire food process - from receiving raw materials to production, labeling, and processing - where allergen risks may occur. It also details the most effective methods for allergen detection. 

Invitek Diagnostics provides a variety of tests capable of detecting target allergens in ingredients, clean-in-place rinses, finished products, and on environmental surfaces. Visit our website to explore our complete portfolio and take a step towards elevating your food safety standards today!

REFERENCES

1. 1. Sharma, G. M., & Pereira, M. (2023). Advances in PCR-based methods for food allergen detection. Journal of Food Protection, 86(3), 456-467. https://doi.org/10.4315/JFP-22-123
   2. Linacero, R., & Cuadrado, C. (2022). New research in food allergen detection. Foods, 11(10), 1520. https://doi.org/10.3390/foods11101520
   3. Sanchiz, A., et al. (2021). Analytical methods for allergen control in food processing. Foods, 10(5), 1120. https://doi.org/10.3390/foods10051120